Haihui Li1, Shanjun Cai2, Xin Gong1, Zhipeng Wu1, Jianping Lyn1, Gang Su1, Bing Xie1. 1. Department of Ophthalmology, Zunyi Medical College Affiliated Hospital, Zunyi 563003, China. 2. Department of Ophthalmology, Zunyi Medical College Affiliated Hospital, Zunyi 563003, China. Email: caishanjun@163.com.
Abstract
OBJECTIVE: To investigate the effect of blue light on human retinal pigment epithelium (RPE) α1D subunit protein expression and its relationship with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) secretion in vitro. METHODS: The fourth generation cultured human RPE cells in vitro were randomly divided into 4 groups, group A (control), group B (exposure to blue light), group C (exposure to blue light+nifedipine), group D [exposure to blue light+(-)Bay K8644]. Cells were exposed to blue light (2 000 ± 500) lx for 6 hours, and cells culture completed 24 hours later. VEGF and bFGF concentration were assayed by enzyme linked immunosorbent assay (ELISA). Real-time polymerase chain reaction was used to analysis L-type calcium channel α1D subunit mRNA expression. Western blot was used to examine the protein expression of L-type calcium channel α1D subunit. Analysis of variance was used to compare the difference of α1D subunit mRNA and protein expression, VEGF and bFGF concentration between groups. Correlation analysis was used to show the relationship between α1D subunit protein expression and concentration of VEGF and bFGF. RESULTS: (1) There is significant statistically difference in the population mean of VEGF and bFGF concentration in four groups (F = 99.441, 21.310, P = 0.000,0.000) . VEGF and bFGF concentration in group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) and group D (3 808.01 ± 94.01, 1 485.82 ± 108.97) was higher than it was in group A (2 401.09 ± 228.07, 1 232.42 ± 65.41) , which was statistically different (P = 0.000, 0.000, 0.019, 0.000). And it was higher in group D (3 808.01 ± 94.01, 1 485.82 ± 108.97) compared with group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) (P = 0.000, 0.006). While, it was lower in group C (1 927.28 ± 143.11, 1 149.39 ± 62.99) than it was in group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) (P = 0.000, 0.000) . (2) The mean of mRNA expression of α1D subunit between four groups was statistically significant (F = 50.320, P = 0.000) . It was higher in group B (210 ± 23) , group D (478 ± 80) and group C (232 ± 14) than group A (100 ± 20). It was statistically significant different (P = 0.023, 0.006, 0.010). And it was higher in group D (478 ± 80) than group B (210 ± 23) and C (232 ± 14) (P = 0.032, 0.039). (3) There was statistically significant difference in the expression of L-type calcium channel α1D subunit protein in four groups (F = 1 940.363, P = 0.000) . It was significantly higher (P = 0.000,0.000) in group B (0.974 2 ± 0.014 7) and group D (0.654 9 ± 0.005 0) than group A (0.503 2 ± 0.007 5). And it was higher in group D (0.654 9 ± 0.005 0) than it was in group C (0.413 9 ± 0.008 8) (P = 0.000). (4) There was positive correlation between the L-type calcium channel α1D subunit protein expression and VEGF concentration secreted by retinal pigment epithelium cells (r = 0.674, F = 8.333, P = 0.016), but there was no correlation with bFGF concentration (r = 0.537, F = 4.061, P = 0.072). CONCLUSIONS: L-type calcium channel α1D subunit may be involved in blue light induced damage on human retinal pigment epithelial cells. Blue light exposure can induce the mRNA and protein expression of α1D subunit, VEGF and bFGF concentration in retinal pigment epithelium cells Increased. And there was a positive correlation between α1D subunit protein expression and the VEGF concentration.
OBJECTIVE: To investigate the effect of blue light on human retinal pigment epithelium (RPE) α1D subunit protein expression and its relationship with vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) secretion in vitro. METHODS: The fourth generation cultured human RPE cells in vitro were randomly divided into 4 groups, group A (control), group B (exposure to blue light), group C (exposure to blue light+nifedipine), group D [exposure to blue light+(-)Bay K8644]. Cells were exposed to blue light (2 000 ± 500) lx for 6 hours, and cells culture completed 24 hours later. VEGF and bFGF concentration were assayed by enzyme linked immunosorbent assay (ELISA). Real-time polymerase chain reaction was used to analysis L-type calcium channel α1D subunit mRNA expression. Western blot was used to examine the protein expression of L-type calcium channel α1D subunit. Analysis of variance was used to compare the difference of α1D subunit mRNA and protein expression, VEGF and bFGF concentration between groups. Correlation analysis was used to show the relationship between α1D subunit protein expression and concentration of VEGF and bFGF. RESULTS: (1) There is significant statistically difference in the population mean of VEGF and bFGF concentration in four groups (F = 99.441, 21.310, P = 0.000,0.000) . VEGF and bFGF concentration in group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) and group D (3 808.01 ± 94.01, 1 485.82 ± 108.97) was higher than it was in group A (2 401.09 ± 228.07, 1 232.42 ± 65.41) , which was statistically different (P = 0.000, 0.000, 0.019, 0.000). And it was higher in group D (3 808.01 ± 94.01, 1 485.82 ± 108.97) compared with group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) (P = 0.000, 0.006). While, it was lower in group C (1 927.28 ± 143.11, 1 149.39 ± 62.99) than it was in group B (3 281.51 ± 251.73, 1 346.81 ± 62.27) (P = 0.000, 0.000) . (2) The mean of mRNA expression of α1D subunit between four groups was statistically significant (F = 50.320, P = 0.000) . It was higher in group B (210 ± 23) , group D (478 ± 80) and group C (232 ± 14) than group A (100 ± 20). It was statistically significant different (P = 0.023, 0.006, 0.010). And it was higher in group D (478 ± 80) than group B (210 ± 23) and C (232 ± 14) (P = 0.032, 0.039). (3) There was statistically significant difference in the expression of L-type calcium channel α1D subunit protein in four groups (F = 1 940.363, P = 0.000) . It was significantly higher (P = 0.000,0.000) in group B (0.974 2 ± 0.014 7) and group D (0.654 9 ± 0.005 0) than group A (0.503 2 ± 0.007 5). And it was higher in group D (0.654 9 ± 0.005 0) than it was in group C (0.413 9 ± 0.008 8) (P = 0.000). (4) There was positive correlation between the L-type calcium channel α1D subunit protein expression and VEGF concentration secreted by retinal pigment epithelium cells (r = 0.674, F = 8.333, P = 0.016), but there was no correlation with bFGF concentration (r = 0.537, F = 4.061, P = 0.072). CONCLUSIONS: L-type calcium channel α1D subunit may be involved in blue light induced damage on human retinal pigment epithelial cells. Blue light exposure can induce the mRNA and protein expression of α1D subunit, VEGF and bFGF concentration in retinal pigment epithelium cells Increased. And there was a positive correlation between α1D subunit protein expression and the VEGF concentration.